Optic nerve-dependent changes in adult frog tectal cell phenotypes

Optic nerve activity helps determine the placement of retinal ganglion cell terminals in the optic tectum of the frog. We investigated whether the presence of this nerve might also influence a characteristic of its target structure, neurotransmitter biosynthesis. We performed unilateral optic nerve transections on adult animals and assayed the percent and intensity of substance P- and serotoninlike immunoreactive (SP-ir and 5-HT-ir, respectively) cells in the deafferented and afferented tectal lobes. Regeneration of the optic nerve was prevented. The percent of SP-ir cells in the afferented tectal lobes was significantly less than that in the deafferented ones either 6 weeks or 5 months following optic nerve lesion. Comparison to normal animals indicated that the change in SP-ir expression was due to a decrease in the percent of immunoreactive cells in the afferented tecta ipsilateral to the optic nerve lesion. The serotoninlike immunoreactivity of tectal cells was also significantly different in the two lobes following optic nerve lesions. This difference resulted from an increase in the percent of 5-HT-ir cells in the deafferented tectum. In addition, the intensity of 5-HT-ir cells in the deafferented lobe was significantly greater than in the afferented one. The staining intensity of SP-ir cells underwent only a transient, relative decrease in the deafferented tectum. We conclude that the optic nerve does regulate substance P and serotonin expression in the tectum, but that this regulation likely occurs through different pathways. © 1996 John Wiley & Sons, Inc.     

The iron regulatory hormone hepcidin reduces ferroportin 1 content and iron release in H9C2 cardiomyocytes

Iron plays a key pathophysiological role in a number of cardiac diseases. Studies on the mechanisms of heart iron homeostasis are therefore crucial for understanding the causes of excessive heart iron. In addition to iron uptake, cellular iron balance in the heart also depends on iron export. We provided evidence for the existence of iron exporter ferroportin 1 (Fpn1) in the heart in a recent study. The presence of hepcidin, a recently discovered iron regulatory hormone, was also confirmed in the heart recently. Based on these findings and the inhibiting role of hepcidin on Fpn1 in other tissues, we speculated that hepcidin might be able to bind with, internalize and degrade Fpn1 and then decrease iron export in heart cells, leading to an abnormal increase in heart iron and iron mediated cell injury. We therefore investigated the effects of hepcidin on the contents of Fpn1 and iron release in H9C2 cardiomyocyte cell line. We demonstrated that hepcidin has the ability to reduce Fpn1 content as well as iron release in this cell. The similar regulation patterns of hepcidin on the Fpn1 and iron release suggested that the decreased iron release resulted from the decreased content of Fpn1 induced by hepcidin. We also found that hepcidin has no significant effects on ceruloplasmin (CP) and hephaestin (Heph) — two proteins required for iron release from mammalian cells. The data imply that Fpn1, rather than Heph and CP, is the limited factor in the regulation of iron release from heart cells under physiological conditions.     

Isolation,Diversity, and Antimicrobial Activity of Rare Actinobacteria from Medicinal Plants of Tropical Rain Forests in Xishuangbanna,China

Endophytic actinobacteria are relatively unexplored as potential sources of novel species and novel natural products for medical and commercial exploitation. Xishuangbanna is recognized throughout the world for its diverse flora, especially the rain forest plants, many of which have indigenous pharmaceutical histories. However, little is known about the endophytic actinobacteria of this tropical area. In this work, we studied the diversity of actinobacteria isolated from medicinal plants collected from tropical rain forests in Xishuangbanna. By the use of different selective isolation media and methods, a total of 2,174 actinobacteria were isolated. Forty-six isolates were selected on the basis of their morphologies on different media and were further characterized by 16S rRNA gene sequencing. The results showed an unexpected level of diversity, with 32 different genera. To our knowledge, this is the first report describing the isolation of Saccharopolyspora, Dietzia, Blastococcus, Dactylosporangium, Promicromonospora, Oerskovia, Actinocorallia, and Jiangella species from endophytic environments. At least 19 isolates are considered novel taxa by our current research. In addition, all 46 isolates were tested for antimicrobial activity and were screened for the presence of genes encoding polyketide synthetases and nonribosomal peptide synthetases. The results confirm that the medicinal plants of Xishuangbanna represent an extremely rich reservoir for the isolation of a significant diversity of actinobacteria, including novel species, that are potential sources for the discovery of biologically active compounds.The class Actinobacteria accounts for a high proportion of soil microbial biomass and contains the most economically significant prokaryotes, producing more than half of the bioactive compounds in a literature survey (46), including antibiotics (6), immunosuppressive agents (55), antitumor agents (18), and enzymes (64). Actinobacteria belonging to the genus Streptomyces, in particular, are excellent producers. The emergence of drug resistance in many bacterial pathogens and the current increase in the number of fungal infections has caused a resurgence of interest in finding new reserves of biologically active compounds (63). As the search for novel natural products continues, it becomes apparent that the rate of discovery of new compounds from soil streptomycetes has decreased, whereas the rate of reisolation of known compounds has increased (28). Recently, evidence has accumulated that rare actinomycete species, which are often very difficult to isolate and cultivate, might represent a unique source of novel biologically active compounds (4). On the other hand, new microbial habitats need to be examined in the search for novel bioactive compounds. One biologically important but relatively overlooked niche is the inner tissues of higher plants. Early studies have demonstrated that some actinobacteria can form intimate associations with plants and colonize their inner tissues. Frankia species and Streptomyces scabies can penetrate their hosts and establish either pathogenic or endophytic associations (5, 24). The actinomycetes that reside in the tissues of living plants and do not visibly harm the plants are known as endophytic actinobacteria (37). These actinobacteria are relatively unstudied and are potential sources of novel natural products for exploitation in medicine, agriculture, and industry (73).Endophytic actinobacteria have attracted attention in recent years, with increasing reports of isolates from a range of plant types, including crop plants (cereals, such as wheat and rice, as well as potatoes, carrots, tomatoes, and citrus) (2, 16, 62, 71, 74, 80) and medicinal plants (75, 88). The culturable endophytic actinobacteria from these plants were found to fall within a narrow species distribution: Streptomyces spp. were the predominant species, and Microbispora, Micromonospora, Nocardioides, Nocardia, and Streptosporangium were the common genera. Endophytic actinobacteria have been demonstrated to improve and promote the growth of host plants as well as to reduce disease symptoms caused by plant pathogens through various mechanisms, including the production of secondary metabolites, which are used in direct antagonism against pests and diseases (9, 10, 12), changes in host physiology (42), and the induction of systemic acquired resistance in plants (15). Another significant function found for these actinobacteria was antibiotic activity, suggesting that endophytic actinobacteria can be an interesting source for bioprospecting. New antibiotics from endophytic Streptomyces spp.—alnumycin, munumbicins A to D, and coronamycins—have been reported (7, 11). Recently, two novel antitumor anthraquinones, lupinacidins A and B, were isolated from a new endophytic Micromonospora sp. (43). Moreover, new species of endophytic actinobacteria have been increasingly reported (25, 35). Thus, endophytic actinobacteria are expected to be potential sources of new species and new bioactive agents.Of the myriad ecosystems on earth, those with the greatest general biodiversity seem also to have the greatest number and the greatest diversity of endophytes (73). Tropical and temperate rain forests are the most biologically diverse terrestrial ecosystems on earth and thus the greatest possible resource for the acquisition of novel microorganisms and their products (73). One area of enormous plant biodiversity is Xishuangbanna, located in the People''s Republic of China at the border with Myanmar. This area lies at the ecotone between the Asian tropics and subtropics and is dominated by tropical seasonal rain forests (87). Xishuangbanna contains more than 5,000 species of vascular plants, comprising 16% of China''s total plant diversity, and more than 3,000 are endemic species (53, 60), many of which have ethnobotanical histories. Until the present, little research was carried out to isolate endophytic actinobacteria and their secondary metabolites from Xishuangbanna (36, 86). In our long-term study of endophytic actinobacterial diversity and bioactive metabolites from tropical rain forest medicinal plants in Xishuangbanna, many bioactive endophytic Streptomyces spp. have been isolated (49). However, the work to date is insufficient to provide a general understanding of the diversity, distribution, and ecology of tropical rain forest endophytic actinobacteria and to facilitate further exploitation of the diverse functions of this novel microbial source.In the present study, the diversity of rare endophytic actinobacteria associated with medicinal plants from the tropical rain forest in Xishuangbanna was investigated by combining special culturing techniques. The selected isolates were also identified by 16S rRNA gene analysis. The overall aims of this study were (i) to analyze the actinobacterial community and reveal whether the rain forest investigated in Xishuangbanna represents a valuable source for abundant endophytic actinobacteria and new species, (ii) to evaluate the antimicrobial activities of these actinobacteria and the biosynthetic potential of related secondary metabolites, and (iii) to study the relationships between the taxa of these endophytic actinobacteria and the isolation methods applied.     

乙型脑炎病毒NS2A-C60A位点突变对其生物学特性影响研究*

目的:旨在探索Ⅰ型日本乙型脑炎病毒传代致弱后基因组突变NS2A-C60A对乙脑病毒生物学特性的影响。方法:首先通过对传代致弱及原始乙脑毒株基因组序列进行测序比对、结构预测分析并利用Western blotting(WB)确定了目标研究位点NS2A-C60A;然后使用反向遗传定点突变技术构建拯救了包含NS2A-C60A单点突变的病毒株;最后利用噬斑形态观察、生长曲线、双萤光素酶分析,WB以及炎性因子检测和动物实验研究了该单点突变对于乙脑病毒生物学特性的影响。结果:首次研究发现Ⅰ型乙脑病毒传代致弱会导致NS1'蛋白表达的显著下降以及可能的相关位点NS2A-C60A,并成功拯救获得了NS2A-C60A单点突变毒株rJEV-C60A,研究发现NS2A-C60A突变对乙脑病毒的生长特性及噬斑形成没有显著影响,但是能够显著降低乙脑病毒NS1'蛋白的表达,并且该位点突变能够轻微阻碍乙脑病毒对细胞炎性因子表达的抑制,动物实验结果显示NS2A-C60A点突变病毒与原毒株具有相似的神经毒力,说明该位点突变不是影响乙脑病毒毒力致弱的关键位点。结论:新发现的NS2A-C60A位点突变能够显著减少乙脑病毒NS1'蛋白的表达,但是对其增殖、诱导炎症及神经毒力等生物学特性没有显著影响。     

Genome Sequence of Staphylococcus capitis QN1, Which Causes Infective Endocarditis

Staphylococcus capitis is a subtype of coagulase-negative staphylococci (CoNS) which could emerge as a significant pathogen causing infective endocarditis, prosthetic valve endocarditis, and late-onset sepsis. We isolated S. capitis strain QN1 from the skin swab sample of a female. Here we prepared a genome sequence for this strain consisting of 30 contigs totaling 2,430,101 bases and a GC content of 32.76%.     

Synthesis and structure–activity relationship studies of phenolic hydroxyl derivatives based on quinoxalinone as aldose reductase inhibitors with antioxidant activity

To enhance aldose reductase (ALR2) inhibition and add antioxidant ability, phenolic hydroxyl was introduced both to the quinoxalinone core and C3 side chain, resulting in a series of derivatives as ALR2 inhibitors. Biological activity tests suggested that most of the derivatives were potent and selective inhibitors with IC₅₀ values ranging from 0.059 to 6.825 μM, and 2-(3-(4-hydroxystyryl)-7-methoxy-2-oxoquinoxalin-1(2H)-yl)acetic acid (6b) was the most active. Particularly, it was encouraging to find that some derivatives endowed with obvious antioxidant activity, and among them the phenolic 3,4-dihydroxyl compound 6f with 7-hydroxyl in the quinoxalinone core showed the most potent activity, even comparable with the well-known antioxidant Trolox. Structure-activity relationship and docking studies highlighted the importance of phenolic hydroxyl both in C3 side chain and the core structure for constructing potent ALR2 inhibitors with antioxidant activity.     

Downregulation of K(+) channel genes expression in type I diabetic cardiomyopathy

Type I diabetic cardiomyopathy has consistently been shown to be associated with decrease of repolarising K(+) currents, but the mechanisms responsible for the decrease are not well defined. We investigated the streptozotocin (STZ) rat model of type I diabetes. We utilized RNase protection assay and Western blot analysis to investigate the message expression and protein density of key cardiac K(+) channel genes in the diabetic rat left ventricular (LV) myocytes. Our results show that message and protein density of Kv2.1, Kv4.2, and Kv4.3 are significantly decreased as early as 14 days following induction of type I diabetes in the rat. The results demonstrate, for the first time, that insulin-deficient type I diabetes is associated with early downregulation of the expression of key cardiac K(+) channel genes that could account for the depression of cardiac K(+) currents, I(to-f) and I(to-s). These represent the main electrophysiological abnormality in diabetic cardiomyopathy and is known to enhance the arrhythmogenecity of the diabetic heart. The findings also extend the extensive list of gene expression regulation by insulin.     

家蚕二分浓核病毒VD1-ORF4基因截短多肽的表达和抗体制备

通过PCR扩增家蚕二分浓核病毒(Bombyx mori bidensovirus,BmBDV)VD1-ORF4基因序列中的两个DNA片段,将测序正确的两个目的片段分别亚克隆到原核表达载体pET-30a上,通过不同浓度的IPTG对含有重组质粒的大肠杆菌BL21(DE3)进行诱导,对诱导产物进行SDS-PAGE和Westen blot分析.结果表明,这两个截短多肽都获得了表达,其N-端融合有6个组氨酸.将割胶纯化的蛋白多肽与佐剂充分研磨,以研磨后的匀浆液对昆明小鼠进行皮下多点注射.获得的抗血清分别对原核诱导表达产物进行Western blot分析,结果表明,在特定的位置都能杂交到一条特异的蛋白带,表明制备的两个多抗能为深入研究VD1-ORF4基因的功能提供基础.